DNA mismatches are classified as either single base mispairs, like G-T, or loops of excess nucleotides on one strand. Mismatches that arise as errors in DNA replication are pre- mutagenic lesions. The well-known mismatch repair (MR) system acts on single base mispairs and on small loops to prevent substitution errors and small insertions and deletions. In the absence of MR, mutation rates are greatly elevated. The ability of MR to correct mismatches therefore provides a powerful mutation avoidance system. Humans with inherited MR deficiencies are prone to hereditary nonpolyposis colon cancer (HNPCC), as well as other types of cancer. Although MR is effective on many mispairs, it does not function on lops larger than about 13 nt. Recent biochemical evidence from yeast and human cells indicates that another repair pathway called large loop repair (LLR) is active on larger heterologies. It is proposed that LLR helps prevent large insertions and deletions by correcting the pre-mutagenic looped mispair. LLR is clearly distinct from MR; for example, LLR is fully active in cells with null mutations in crucial MR genes. The objectives of this project are to deduce key biochemical aspects of yeast LLR, to identify required proteins, and to begin establishing the importance of LLR in genetic stabilization. Two complementary approaches are proposed that utilize highly specific biochemical and genetic assays developed in this laboratory. This project will help advance the field of DNA repair by characterizing a novel, conserved, eukaryote-specific repair pathway. The research will also reveal new DNA repair proteins and provide information on their biochemical functions. This project will also address the genetic impact of LLR. Yeast was chosen because loop repair has been demonstrated in this organism, because it provides a simple eukaryotic system and because the experimental tractability of yeast allows one to answer important questions about loop repair. Based on the precedent that basic science approaches to MR in yeast led to the connection with human HNPCC, it is suggested that studies of yeast LLR may also help identify the mutational cause of other hereditary cancers or genetic diseases.